The concept of intracellular immunization - insertion of a gene into a cell to express a product which confers resistance to infection of the cell by a pathogen - has been investigated as a potential therapeutic approach to HIV-1 infection. A large number of synthetic genes have been developed which can inhibit infection or replication of HIV-1 ("anti HIV-1 genes"). These anti-HIV-I genes are able to strongly suppress HIV-1 replication when tested in human cell lines and primary cells in culture. Hematopoietic stem cells (HSC) that reside in the bone marrow are a theoretically better target for anti-HIV-I genes than mature T lymphocytes for a number of reasons. HSC give rise to all of the cells involved in HIV-1 pathogenesis (CD4 T lymphocytes, monocytes, dendritic cells, macrophages, microglia) and thus effective transduction of HSC with anti-HIV-1 genes could give rise to gene-protected cells of all the key lineages. While effective and non- toxic anti-HIV-1 genes have been developed, clinical trials performed to date using oncoretroviral vectors have not achieved transduction and engraftment of long-term HSC. Lentiviral vectors have been developed and may be more effective at transduction of HSC than oncoretroviral vectors. Busulfan is a chemotherapeutic agent that produces a specific elimination of HSC and has been used extensively as a cytoreductive conditioning agent prior to bone marrow transplantation (BMT). The central hypothesis of this proposal is that lentiviral vectors expressing anti-HIV-1 genes will be able to effectively transduce HSC and these HSC will have augmented engraftment in autologous transplantation using an optimal cytoreductive dosage of busulfan. We will use an infant rhesus monkey model to evaluate whether lentiviral vector transduction of HSC and cytoreduction with busulfan will lead to engraftment of gene-containing HSC and whether transduction of HSC with an anti-HIV-1 gene and engraftment augmented with busulfan will confer a selective survival advantage on peripheral blood T lymphocytes, suppress SHIV replication and slow the progression of SHIV-induced disease. Overall, the studies of this proposal will advance the potential for applying gene therapy as a new strategy for the treatment of HIV-1 infection. This "intracellular immunization" approach may provide a therapeutic measure against HIV-1 that adds to pre-existing therapies, such as anti-retroviral drugs.